This invention relates broadly to novel thiazolidinones. More specifically, the invention relates to thiazolidinones which modulate Follicle Stimulating Hormone (FSH) activity.
Approximately 400,000 germ cells are stored in the ovaries of the human female at the time of puberty. No further germ cells are made. Beginning at the time of puberty and ending at menopause, there are approximately 400 ovulatory menstrual cycles which consume essentially all of the germ cells in the human ovary. About 1,000 germ cells are consumed in each menstrual period. However, in any one menstrual cycle, only one germ cell, developed in what becomes the dominant follicle, is ovulated and available for pregnancy.
Although the details are not accurately known, the mechanism by which a single egg is selected each month to become the dominant egg is dependent upon a complex interaction between one or more hormones from the ovary, hypothalamus and the pituitary. Three glycoprotein hormones (luteinizing hormone (LH), follicle stimulating hormone (FSH) and chorionic gonadotropin (hCG)) act on the ovary to stimulate steroid synthesis and secretion. LH and FSH are secreted by the pituitary and together play a central role in regulating the menstrual cycle and ovulation. hCG is secreted by the developing placenta from the early stages of pregnancy and its role is to maintain steroid secretion by the corpus luteum, which is necessary to prevent ovulation during pregnancy.
In the normal cycle, there is a mid-cycle surge in LH concentration which is followed by ovulation. An elevated estrogen level, which is brought about by the endogenous secretion of LH and FSH, is required for the LH surge to occur. The estrogen mediates a positive feedback mechanism which results in the increased LH secretion.
Oral contraceptive agents have been used by over 200 million women worldwide and by 1 of 4 women in the United States under the age of 45. Such agents are popular because of ease of administration, low pregnancy rate (less than 1 percent) and a relatively low incidence of side effects. Typically, oral contraceptives inhibit ovulation by suppressing FSH and LH secretion. As a consequence, the secretion of all ovarian steroids is also suppressed, including estrogen, progesterone and androgen. These agents also exert minor direct inhibitory effects on the reproductive tract, altering the cervical mucus, thereby decreasing sperm penetration and decreasing the motility and secretions of the fallopian tubes and uterus.
Thiazolidinones are a class of small molecule organic compounds which have found limited pharmaceutical use. For example, thiazolidinones have been found to have central nervous system activity. See, for example, Tripathi, et al., xe2x80x9cThiazolidinone congeners as central nervous system active agents.xe2x80x9d Arzneimittelforschung 43:632-5 (1993). CNS activities which have been identified include, for example, antipsychotic properties. See, Mutlib, et al., xe2x80x9cMetabolism of an atypical antipsychotic agent, 3-[4-[4-(6-fluorobenzo[b]thien-3-yl)-1-piperazinyl]butyl]-2,5,5-trimethyl-4-thiazolidinone (HP236).xe2x80x9d Drug Metab. Dispos. 24:1139-50 (1996). Other thiazolidinones have been found to be CNS antiischemic agents. See, Ruterbories, et al., xe2x80x9cPharmacokinetics of a novel butylated hydroxytoluene-thiazolidinone CNS antiischemic agent LY256548 in rats, mice, dogs and monkeys.xe2x80x9d Drag Metab. Dispos. 18:674-9 (1990). Thiazolidinones have also been used as antimicrobial agents. See, for example, Ley, et al., xe2x80x9cInhibition of multiplication of Mycobacterium leprae by several antithyroid drugs.xe2x80x9d Am. Rev. Respir. Dis. 111:651-5 (1975).
The synthesis of novel thiazolidinones offers the promise for discovering new pharmaceutical agents with applications in areas as diverse as, for example, antimicrobial therapy and the treatment of strokes with CNS antiischemic agents. Of particular interest is the use of novel thiazolidinones as regulators of mammalian fertility.
Although a number of oral contraceptives are commercially available, there still remains a need for new fertility-regulating agents which are useful for both in vivo and in vitro applications. A class of small molecule FSH receptor antagonist compounds which are inexpensive to prepare, easily purified, easily administered and which exhibit a broad range of activities would represent a significant advance in the field of oral contraceptive agents. Quite surprisingly, the present invention provides such small molecule thiazolidinone FSH receptor antagonists.
The present invention provides a class of novel thiazolidinones possessing a range of pharmaceutical applications and activities. Thus, in one aspect, the present invention provides novel thiazolidinones having the formula: 
wherein,
R1 is a member selected from the group consisting of aryl and substituted aryl, alkyl and substituted alkyl;
R2 is a member selected from the group consisting of heterocyclic and substituted heterocylic groups;
R3 and R4 are independently members selected from the group including hydrogen and xe2x80x94(CH2)mCONR5R6;
R5 and R6 are independently members selected from the group consisting of hydrogen, alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, arylalkyl, substituted arylalkyl, heterocyclicalkyl and substituted heterocyclicalkyl groups;
X is a member selected from the group consisting of S, Sxe2x95x90O, and Oxe2x95x90Sxe2x95x90O;
m is a number from 0 to 3.
In a second aspect, the present invention provides novel thiazolidinones 
wherein,
R1 is a member selected from the group consisting of aryl and substituted aryl alkyl and substituted alkyl;
R21, R22, and R23 are members independently selected from the group consisting of H, halogen, lower alkyl, substituted lower alkyl, lower alkoxy, substituted lower alkoxy, phenyl, substituted phenyl, aryloxy, substituted aryloxy, alkynyl, substituted alkynyl and nitro groups. Preferred aryloxy groups are phenoxy and benzyloxy and preferred substituted aryloxy groups are substituted phenoxy and substituted benzyloxy.
Y is a member selected from the group consisting of xe2x80x94Oxe2x80x94, xe2x80x94Sxe2x80x94 and NR24 wherein R24 is H or lower alkyl.
R5 and R6 are independently members selected from the group consisting of hydrogen, alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, arylalkyl, substituted arylalkyl, heterocyclicalkyl and substituted heterocyclicalkyl groups; and
X is a member selected from the group consisting of S, Sxe2x95x90O, and Oxe2x95x90Sxe2x95x90O.
In another aspect, the invention provides a class of FSH receptor antagonists, wherein the receptor antagonists are noncompetitve with FSH for the receptor FSH binding site.
In yet another aspect, the invention provides a class of compounds that modulate FSH hormone activity, the compounds having: (a) a molecular weight of from about 200 daltons to about 1000 daltons; and (b) an FSH antagonist activity corresponding to an IC50 standard of no more than 25 xcexcM, preferably no more than 11 xcexcM; wherein the antagonist activity of this class of compounds to the FSH receptor is competitively inhibited by a compound described above.
In a preferred embodiment, this class of compounds has a molecular weight of about 300 daltons to about 800 daltons. In another preferred embodiment, this class of compounds has an FSH receptor antagonist activity, as expressed by an IC50 standard, of no more than 11 xcexcM.
In still another aspect, the invention provides methods of using the compounds, i.e., thiazolidinones, for diverse pharmaceutical applications including, for example, CNS antiischemic agents, agents with antipsychotic or other psychoactive properties, antimicrobial agents and mammalian fertility regulating agents. When used as mammalian fertility regulating agents, the thiazolidinones are preferably antagonists of the FSH receptor.
As such, in another aspect, the present invention provides pharmaceutical compositions which contain one or more of the compounds of the invention in conjunction with pharmaceutically acceptable excipients, carriers, diluents, etc. The pharmaceutical compositions can also contain agents which are themselves pharmacologically active and which serve to enhance, supplement, decrease or otherwise regulate the pharmacological effect of the pharmaceutical compositions.
Other features, objects and advantages of the invention and its preferred embodiments will become apparent from the detailed description which follows.
HATU, [O-(7-Azabenzotriazol-1-yl)-1,1,3,3-tetramethyluroniumhexafluorophosphate]; DIEA, diisopropylethylamine; FMOC, fluorenylmethoxycarbonyl; DECP, diethyl cyanophosphonate; DCM, dichloromethane; DBU, 1,8-diazabicyclo[5.4.0]undec-7-ene; CHO, chinese hamster ovary; RBF, round-bottomed flask.
The term xe2x80x9cindependently selectedxe2x80x9d is used herein to indicate that the R groups, e.g., R1, R2, and R3, can be identical or different (e.g., R1, R2 and R3 may all be substituted alkyls or R1 and R2 may be a substituted alkyl and R3 may be an aryl, etc.).
A named R group will generally have the structure which is recognized in the art as corresponding to R groups having that name. For the purposes of illustration, representative R groups as enumerated above are defined herein. These definitions are intended to supplement and illustrate, not preclude, the definitions known to those of skill in the art.
The term xe2x80x9calkylxe2x80x9d is used herein to refer to a branched or unbranched, saturated or unsaturated, monovalent hydrocarbon radical having from 1-12 carbons and preferably, from 1-6 carbons. When xe2x80x9clower alkylxe2x80x9d is used, it refers to an alkyl group which has from 1-6 carbons. Suitable alkyl radicals include, for example, methyl, ethyl, n-propyl, i-propyl, 2-propenyl (or allyl), n-butyl, t-butyl (or 2-methylpropyl), etc.
xe2x80x9cSubstituted alkylxe2x80x9d refers to alkyl as just described including one or more functional groups such as lower alkyl, aryl, acyl, halogen, (i.e., alkylhalos, e.g., CF3), hydroxy, nitro, cyano, amino, alkoxy, alkylamino, acylamino, acyloxy, aryloxy, aryloxyalkyl, mercapto, carboxylic acid, carboxylic acid derivatives, carboxylic acid amides, sulfonic acids, sulfonic acid derivatives, both saturated and unsaturated cyclic hydrocarbons, heterocycles and the like. These groups may be attached to any carbon of the alkyl moiety.
The term xe2x80x9carylxe2x80x9d is used herein to refer to an aromatic substituent having a single aromatic ring or multiple aromatic rings which are fused together, linked covalently, or linked to a common group such as a methylene or ethylene moiety. The common linking group may also be a carbonyl as in benzophenone. The aromatic ring(s) may include phenyl, naphthyl, biphenyl, diphenylmethyl and benzophenone among others.
xe2x80x9cSubstituted arylxe2x80x9d refers to aryl as just described including one or more functional groups such as lower alkyl, acyl, halogen, alkylhalos (e.g., CF3), hydroxy, nitro, cyano, amino, alkoxy, alkylamino, acylamino, acyloxy, mercapto, carboxylic acid amide, sulfonic acid amide and both saturated and unsaturated cyclic hydrocarbons which are fused to the aromatic ring(s), linked covalently or linked to a common group such as a methylene or ethylene moiety. The linking group may also be a carbonyl such as in cyclohexyl phenyl ketone.
The term xe2x80x9carylalkylxe2x80x9d is used herein to refer to a subset of xe2x80x9carylxe2x80x9d in which the aryl group is attached through an alkyl group as defined herein. Examples include, but are not limited to, benzyl, phenylethyl and phenylpropyl groups.
xe2x80x9cSubstituted arylalkylxe2x80x9d defines a subset of xe2x80x9carylalkylxe2x80x9d wherein the aryl moiety of the arylalkyl group is substituted as defined herein for aryl groups.
The term xe2x80x9chalogenxe2x80x9d is used herein to refer to fluorine, bromine, chlorine and iodine atoms.
The term xe2x80x9chydroxyxe2x80x9d is used herein to refer to the group xe2x80x94OH.
The term xe2x80x9caminoxe2x80x9d is used herein to refer to the group-NRRxe2x80x2, where R and Rxe2x80x2 may independently be hydrogen, lower alkyl, substituted lower alkyl, aryl, substituted aryl or acyl.
The term xe2x80x9calkoxyxe2x80x9d is used herein to refer to the xe2x80x94OR group, where R is a lower alkyl or substituted lower alkyl, wherein the alkyl and substituted lower alkyl groups are as described herein. Suitable alkoxy radicals include, for example, methoxy, ethoxy, t-butoxy, etc.
The term xe2x80x9caryloxyxe2x80x9d is used herein to refer to the xe2x80x94OR group, wherein R is an aryl, substituted aryl, arylalkyl or substituted arylalkyl as described above. Examples include phenoxy, benzyloxy, phenethyloxy and substituted derivatives thereof.
The term xe2x80x9calkylaminoxe2x80x9d denotes secondary and tertiary amines wherein the alkyl groups may be either the same or different and may consist of straight or branches, saturated or unsaturated hydrocarbons.
The term xe2x80x9cheterocyclicxe2x80x9d is used herein to describe a monovalent group having a single ring or multiple condensed rings from 1-12 carbon atoms and from 1-4 heteroatoms selected from nitrogen, sulfur or oxygen within the ring. Heterocyclic groups include saturated, unsaturated, and aromatic ring systems. Such heterocycles include, for example, tetrahydrofuran, morpholine, piperidine, pyrrolidine, thiophene, pyridine, isoxazole, phthalimide, pyrazole, indole, furan, benzo-fused analogs of these rings, etc.
The term xe2x80x9csubstituted heterocyclicxe2x80x9d as used herein describes a subset of xe2x80x9cheterocyclicxe2x80x9d wherein the heterocycle nucleus is substituted with one or more functional groups such as lower alkyl, acyl, halogen, alkylhalos (e.g., CF3), hydroxy, amino, alkoxy, alkylmino, acylamino, acyloxy, mercapto, etc.
The term xe2x80x9cheterocyclicalkylxe2x80x9d is used herein to refer to a subset of xe2x80x9cheterocylicxe2x80x9d in which the hetetocylcic group is attached through an alkyl group as defined herein.
xe2x80x9cSubstituted heterocyclicalkylxe2x80x9d defines a subset of xe2x80x9cheterocyclicalkylxe2x80x9d wherein the heterocyclic moiety of the hetemocyclicalkyl group is substituted as defined herein for heterocyclic groups.
The term xe2x80x9cpharmaceutically acceptable saltxe2x80x9d refers to those salts of compounds which retain the biological effectiveness and properties of the free bases and which are obtained by reaction with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid and organic acids such as, for example, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid and the like. Pharmaceutically acceptable salts include, for example, alkali metal salts, such as sodium and potassium, alkaline earth salts and ammonium salts.
The term xe2x80x9ccontactingxe2x80x9d is used herein interchangeably with the following: combined with, added to, mixed with, passed over, incubated with, flowed over, etc. Moreover, the thiazolidinone compounds of present invention can be xe2x80x9cadministeredxe2x80x9d to a subject by any conventional method such as, for example, parenteral, oral, topical and inhalation routes as described herein.
xe2x80x9cAn amount sufficientxe2x80x9d or xe2x80x9can effective amountxe2x80x9d is that amount of a given thiazolidinone analog which exhibits the binding/activity of interest or, which provides an improvement in gamete recruitment.
xe2x80x9cIC50xe2x80x9d is the inhibitory concentration: the concentration of a compound at which 50% of the maximal response of that obtained with FSH is inhibited.
xe2x80x9cNon-competitivexe2x80x9d refers to the nature of the agonist activity exhibited by the compounds of the invention, wherein the compounds act as agonists of and activate the FSH receptor without substantially reducing the magnitude of binding of FSH to the receptor. xe2x80x9cMagnitude of bindingxe2x80x9d refers to the amount of FSH bound by a receptor population and/or the strength of the binding interaction between FSH and the FSH receptor.
The present invention is directed to novel thiazolidinone compounds which exhibit a range of pharmaceutical activities. In a presently preferred embodiment, the novel compounds are small molecule FSH receptor antagonists. These compounds offer numerous advantages over the current state of the art. For example, the compounds of the instant invention are inexpensive and both easily prepared and purified. Further, the compounds exhibit a range of activity regarding the FSH receptor. Such a manifold of compounds of differing activity provides an opportunity to the clinician to modulate the desired level of fertility induction by judicious choice of the fertility-inhibiting agent. In addition, the novel thiazolidinones, as small molecules, exhibit a pharmacokinetic profile which is distinct from that of conventional peptidic hormone preparations. The pharmacokinetic profile can be further modified by judicious choice of the route of administration and manipulating the nature of the substituents on the thiazolidinone nucleus.
As such, in a first aspect, the present invention provides novel thiazolidinones having the formula: 
wherein,
R1 is a member selected from the group consisting of aryl and substituted aryl, alkyl and substituted alkyl;
R2 is a member selected from the group consisting of heterocyclic and substituted heterocylic groups;
R3 and R4 are independently members selected from the group including hydrogen and xe2x80x94(CH2)mCONR5R6;
R5 and R6 are independently members selected from the group consisting of hydrogen, alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, arylalkyl, substituted arylalkyl, heterocyclicalkyl and substituted heterocyclicalkyl groups;
X is a member selected from the group consisting of S, Sxe2x95x90O, and Oxe2x95x90Sxe2x95x90O;
m is a number from 0 to 3.
A more preferred embodiment of Formula (I), are novel thiazolidinones of Formula (III) 
wherein,
R1 is a member selected from the group consisting of aryl and substituted aryl alkyl and substituted alkyl;
R21, R22, and R23 are members independently selected from the group consisting of H, halogen, lower alkyl, substituted lower alkyl, lower alkoxy, substituted lower alkoxy, phenyl, substituted phenyl, aryloxy, substituted aryloxy, alkynyl, substituted alkynyl and nitro groups. Preferred aryloxy groups are phenoxy and benzyloxy and preferred substituted aryloxy groups are substituted phenoxy and substituted benzyloxy.
Y is a member selected from the group consisting of xe2x80x94Oxe2x80x94, xe2x80x94Sxe2x80x94 and NR24 wherein R24 is H or lower alkyl
R5 and R6 are independently members selected from the group consisting of hydrogen, alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, arylalkyl, substituted arylalkyl, heterocyclicalkyl and substituted heterocyclicalkyl groups; and
X is a member selected from the group consisting of S, Sxe2x95x90O, and Oxe2x95x90Sxe2x95x90O.
In yet a further preferred embodiment, the present invention provides a compound wherein, when substituent R4, on C-5, is H, and a second substituent at C-5 (R3) is not H, said substituent R3 on C-5 and substituent R2 on C-2 are oriented in a cis manner.
In certain presently preferred embodiments, R21, R22 and R23 are independently chosen from hydrogen and the groups according to Formulae (VIII): 
wherein R41, R42, R43, R44 and R45 are members independently selected from the group consisting of hydrogen, halogen, nitro and trifluoromethyl.
Due to the chiral carbons at positions 2 and 5 (i.e., C-2 and C-5) of the thiazolidinone ring structure, the compounds of the invention can exist in a number of different isomeric and stereoisomeric forms. The configuration of C-2 and C-5 can be such that their substituents are in either a cis or traits configuration. In preferred embodiments, the compounds exist in the cis configuration. Additionally, the combination of absolute configurations available to C-2 and C-5 can take any one of four permutations. Thus, the thiazolidinone nucleus can be 2S, 5S; 2R, 5R; 2S, SR; or 2R, 5S. Presently preferred embodiments are those in which the configuration at C-2 and C-5 are 2S, 5R.
The compounds of the present invention can be used for diverse pharmaceutical applications including, for example, CNS antiischemic agents, agents with antipsychotic or other psychoactive properties, antimicrobial agents and mammalian fertility regulating agents. When used as mammalian fertility regulating agents, the thiazolidinones are preferably antagonists of the FSH receptor.
Examples of the thiazolidinone compounds of the present invention are displayed below. In a preferred embodiment, the IC50 values of the compounds having antagonist activity are less than 25 xcexcM. In a more preferred embodiment, the IC50 values of the compounds having antagonist activity are less than 11 xcexcM. 
In another aspect, the invention provides a class of FSH receptor antagonists, wherein the receptor antagonist activity is noncompetitve with FSH. In a preferred embodiment, the non-competitive FSH antagonists are organic molecules with a molecular weight of from about 200 daltons to about 1000 daltons. In another preferred embodiment, the invention provides for pharmaceutical formulations containing a FSH receptor antagonist which is non-competitive with FSH. In this aspect, the invention provides regulators of mammalian fertility which are useful in the diverse applications described herein for the thiazolidinones of the invention.
In another embodiment, the present invention provides pharmaceutical compositions which contain one or more of the compounds of the invention in conjunction with pharmaceutically acceptable excipients, carriers, diluents, etc. The pharmaceutical compositions can also contain other agents which are themselves pharmacologically active and which serve to enhance, supplement, decrease or otherwise regulate the pharmacological effect of the pharmaceutical compositions.
The compounds, i.e., thiazolidinones, of the present invention can be administered to a mammal, e.g., a human patient, alone, in the form of a pharmaceutically acceptable salt, or in the form of a pharmaceutical composition where the compound is mixed with suitable carriers or excipient(s) in a therapeutically effective amount. Further, the compounds and compositions of the invention can be administered to induce menses; to treat endometriosis; to treat dysmenorrhea; to treat endocrine hormone-dependent tumors; to treat uterine fibroids; to inhibit uterine endometrial proliferation; to induce labor; for hormone therapy; and for contraception in both the female and male.
More particularly, the compounds of the present invention are of particular value in the control of hormonal irregularities in the menstrual cycle, for controlling endometriosis and dysmenorrhea, and for inducing menses. In addition, the compounds of the present invention can be used as a method of providing hormone therapy either alone or in combination with estrogenic substances in postmenopausal women, or in women whose ovarian hormone production is otherwise compromised.
Moreover, the compounds of the present invention can be used for control of fertility during the whole of the reproductive cycle. For long-term contraception, the compounds of the present invention can be administered either continuously or periodically depending on the dose. In addition, the compounds of the present invention are of particular value as postcoital contraceptives, for rendering the uterus inimical to implantation, and as xe2x80x9conce a monthxe2x80x9d contraceptive agents. They can be used in conjunction with prostaglandins, oxytocics and the like.
A further important utility for the compounds of the present invention lies in their ability to slow down growth of hormone-dependent cancers. Such cancers include kidney, breast, endometrial, ovarian cancers, and prostate cancer which are characterized by possessing progesterone receptors and can be expected to respond to the products of this invention. Other utilities of the compounds of the present invention include the treatment of fibrocystic disease of the breast and uterine.
By analogy to the demonstrated efficacy of gonadotrophins on the Sertoli cell, that is, the male equivalent of the ovarian granulosa cells, the compounds and compositions of the present invention can be used for male, as well as female, contraception. See, for example, Reichert, et al., xe2x80x9cThe follicle stimulating hormone (FSH) receptor in testis: interaction with FSH, mechanism of signal transduction, and properties of the purified receptor,xe2x80x9d Biol. Reprod. 40:13-26 (1989), the disclosure of which is incorporated herein by reference.
The compounds of this invention can be incorporated into a variety of formulations for therapeutic administration. More particularly, the compounds of the present invention can be formulated into pharmaceutical compositions by combination with appropriate, pharmaceutically acceptable carriers or diluents, and may be formulated into preparations in solid, semi-solid, liquid or gaseous forms, such as tablets, capsules, pills, powders, granules, dragees, gels, slurries, ointments, solutions, suppositories, injections, inhalants and aerosols. As such, administration of the compounds can be achieved in various ways, including oral, buccal, rectal, parenteral, intraperitoneal, intradermal, transdermal, intracheal, etc., administration. Moreover, the compound can be administered in a local rather than systemic manner, for example, via injection of the compound directly into an ovary, often in a depot or sustained release formulation. In addition, the compounds can be administered in a targeted drug delivery system, for example, in a liposome coated with an organ surface receptor-specific antibody. Such liposomes will be targeted to and taken up selectively by the organ.
In pharmaceutical dosage forms, the compounds may be administered in the form of their pharmaceutically acceptable salts, or they may also be used alone or in appropriate association, as well as in combination with other pharmaceutically active compounds. In the interest of brevity, the discussion which follows is based on the use of the compounds of the invention as contraceptive agents. That pharmaceutical compositions containing the novel thiazolidinones are useful in other applications, and are not limited to use as contraceptive agents will be apparent to those of skill in the art. In these further applications, adjuncts which serve a purpose analogous to those discussed below (i.e., enhance or supplement the thiazolidinone therapeutic activity) can be included within the formulation.
The thiazolidinone analogs of the present invention can be administered alone, in combination with each other, or they can be used in combination with other known compounds (e.g., contraceptive agent). A number of suitable formulations for use in the present invention are found in Remington""s Pharmaceutical Sciences (Mack Publishing Company, Philadelphia, Pa., 17th ed. (1985), which is incorporated herein by reference. Moreover, for a brief review of methods for drug delivery, .ee, Langer, Science 249:1527-1533 (1990), which is incorporated herein by reference. The pharmaceutical compositions described herein can be manufactured in a manner that is known to those of skill in the art, i.e., by means of conventional mixing, dissolving, granulating, dragee-making, levigating, emulsifying, encapsulating, entrapping or lyophilizing processes. The following methods and excipients are merely exemplary and are in no way limiting.
For injection, the compounds can be formulated into preparations by dissolving, suspending or emulsifying them in an aqueous or nonaqueous solvent, such as vegetable or other similar oils, synthetic aliphatic acid glycerides, esters of higher aliphatic acids or propylene glycol; and if desired, with conventional additives such as solubilizers, isotonic agents, suspending agents, emulsifying agents, stabilizers and preservatives. Preferably, the compounds of the invention may be formulated in aqueous solutions, preferably in physiologically compatible buffers such as Hanks""s solution, Ringer""s solution, or physiological saline buffer. For transmucosal administration, penetrants appropriate to the barrier to be permeated are used in the formulation. Such penetrants are generally known in the art.
For oral administration, the compounds can be formulated readily by combining with pharmaceutically acceptable carriers that are well known in the art. Such carriers enable the compounds to be formulated as tablets, pills, dragees, capsules, emulsions, lipophilic and hydrophilic suspensions, liquids, gels, syrups, slurries, suspensions and the like, for oral ingestion by a patient to be treated. Pharmaceutical preparations for oral use can be obtained by mixing the compounds with a solid excipient, optionally grinding a resulting mixture, and processing the mixture of granules, after adding suitable auxiliaries, if desired, to obtain tablets or dragee cores. Suitable excipients are, in particular, fillers such as sugars, including lactose, sucrose, mannitol, or sorbitol; cellulose preparations such as, for example, maize starch, wheat starch, rice starch, potato starch, gelatin, gum tragacanth, methyl cellulose, hydroxypropylmethyl-cellulose, sodium carboxymethylcellulose, and/or polyvinylpyrrolidone (PVP). If desired, disintegrating agents may be added, such as the cross-linked polyvinyl pyrrolidone, agar, or alginic acid or a salt thereof such as sodium alginate.
Dragee cores are provided with suitable coatings. For this purpose, concentrated sugar solutions may be used, which may optionally contain gum arabic, talc, polyvinyl pyrrolidone, carbopol gel, polyethylene glycol, and/or titanium dioxide, lacquer solutions, and suitable organic solvents or solvent mixtures. Dyestuffs or pigments may be added to the tablets or dragee coatings for identification or to characterize different combinations of active compound doses.
Pharmaceutical preparations which can be used orally include push-fit capsules made of gelatin, as well as soft, sealed capsules made of gelatin and a plasticizer, such as glycerol or sorbitol. The push-fit capsules can contain the active ingredients in admixture with filler such as lactose, binders such as starches, and/or lubricants such as talc or magnesium stearate and, optionally, stabilizers. In soft capsules, the active compounds may be dissolved or suspended in suitable liquids, such as fatty oils, liquid paraffin, or liquid polyethylene glycols. In addition, stabilizers may be added. All formulations for oral administration should be in dosages suitable for such administration.
For buccal administration, the compositions may take the form of tablets or lozenges formulated in conventional manner.
For administration by inhalation, the compounds for use according to the present invention are conveniently delivered in the form of an aerosol spray presentation from pressurized packs or a nebulizer, with the use of a suitable propellant, e.g., dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gas, or from propellant-free, dry-powder inhalers. In the case of a pressurized aerosol the dosage unit may be determined by providing a valve to deliver a metered amount. Capsules and cartridges of, e.g., gelatin for use in an inhaler or insufflator may be formulated containing a powder mix of the compound and a suitable powder base such as lactose or starch.
The compounds may be formulated for parenteral administration by injection, e.g., by bolus injection or continuous infusion. Formulations for injection may be presented in unit dosage form, e.g., in ampules or in multidose containers, with an added preservative. The compositions may take such forms as suspensions, solutions or emulsions in oily or aqueous vehicles, and may contain formulatory agents such as suspending, stabilizing and/or dispersing agents.
Pharmaceutical formulations for parenteral administration include aqueous solutions of the active compounds in water-soluble form. Additionally, suspensions of the active compounds may be prepared as appropriate oily injection suspensions. Suitable lipophilic solvents or vehicles include fatty oils such as sesame oil, or synthetic fatty acid esters, such as ethyl oleate or triglycerides, or liposomes. Aqueous injection suspensions may contain substances which increase the viscosity of the suspension, such as sodium carboxymethyl cellulose, sorbitol, or dextran. Optionally, the suspension may also contain suitable stabilizers or agents which increase the solubility of the compounds to allow for the preparation of highly concentrated solutions. Alternatively, the active ingredient may be in powder form for constitution with a suitable vehicle, e.g., sterile pyrogen-free water, before use.
The compounds may also be formulated in rectal compositions such as suppositories or retention enemas, e.g., containing conventional suppository bases such as cocoa butter, carbowaxes, polyethylene glycols or other glycerides, all of which melt at body temperature, yet are solidified at room temperature.
In addition to the formulations described previously, the compounds may also be formulated as a depot preparation. Such long acting formulations may be administered by implantation (for example subcutaneously or intramuscularly) or by intramuscular injection. Thus, for example, the compounds may be formulated with suitable polymeric or hydrophobic materials (for example as an emulsion in an acceptable oil) or ion exchange resins, or as sparingly soluble derivatives, for example, as a sparingly soluble salt.
Alternatively, other delivery systems for hydrophobic pharmaceutical compounds may be employed. Liposomes and emulsions are well known examples of delivery vehicles or carriers for hydrophobic drugs. Certain organic solvents such as dimethylsulfoxide also may be employed, although usually at the cost of greater toxicity. Additionally, the compounds may be delivered using a sustained-release system, such as semipermeable matrices of solid hydrophobic polymers containing the therapeutic agent. Various types of sustained-release materials have been established and are well known by those skilled in the art. Sustained-release capsules may, depending on their chemical nature, release the compounds for a few weeks up to over 100 days.
The pharmaceutical compositions also may comprise suitable solid or gel phase carriers or excipients. Examples of such carriers or excipients include but are not limited to calcium carbonate, calcium phosphate, various sugars, starches, cellulose derivatives, gelatin, and polymers such as polyethylene glycols.
Pharmaceutical compositions suitable for use in the present invention include compositions wherein the active ingredients are contained in a therapeutically effective amount. The amount of composition administered will, of course, be dependent on the subject being treated, on the subject""s weight, the severity of the affliction, the manner of administration and the judgment of the prescribing physician. Determination of an effective amount is well within the capability of those skilled in the art, especially in light of the detailed disclosure provided herein.
For any compound used in the method of the invention, a therapeutically effective dose can be estimated initially from cell culture assays. For example, a dose can be formulated in animal models to achieve a circulating concentration range that includes the EC50 as determined in cell culture. Such information can be used to more accurately determine useful doses in humans. Initial dosages can also be estimated from in vitro or in vivo data.
Initial dosages can also be formulated by comparing the effectiveness of the compounds described herein in cell culture assays with the effectiveness of known drugs. For instance, when used as contraceptive agents, initial dosages can be formulated by comparing the effectiveness of the compounds described herein in cell culture assays with the effectiveness of known contraceptive agents. In this method, an initial dosage can be obtained by multiplying the ratio of effective concentrations obtained in cell culture assay for the compound of the present invention and a known contraceptive drug by the effective dosage of the known contraceptive drug. For example, if a compound of the present invention is twice as effective in cell culture assay as the known contraceptive agent (i.e., the IC50 of that compound is equal to one-half the IC50 of the known contraceptive agent in the same assay), an initial effective dosage of the compound of the present invention would be one-half the known dosage for the known contraceptive agent. Using these initial guidelines one having ordinary skill in the art could determine an effective dosage in humans or other mammals.
Moreover, toxicity and therapeutic efficacy of the compounds described herein can be determined by standard pharmaceutical procedures in cell cultures or experimental animals, e.g., by determining the LD50 (the dose required to cause death in 50% of the subjects tested) and the ED50 (the dose that produces a defined effect in 50% of the subjects tested). The dose ratio between toxic and therapeutic effect is the therapeutic index and can be expressed as the ratio between LD50 and ED50. Compounds which exhibit high therapeutic indices are preferred. The data obtained from these cell culture assays and animal studies can be used in formulating a dosage range that is appropriate for use in humans. The dosage of such compounds lies preferably within a range of circulating concentrations that include the ED50 with little or no toxicity. The dosage may vary within this range depending upon the dosage form employed and the route of administration utilized. The exact formulation, route of administration and dosage can be chosen by the individual physician in view of the patient""s condition. See, for example, Fingl, et al., In: The Pharmacological Basis of Therapeutics, Ch. 1, p. 1 (1975).
Dosage amount and interval may be adjusted individually to provide plasma levels of the active compound which are sufficient to maintain therapeutic effect. Usual patient dosages for oral administration range from about 50-2000 mg/kg/day, commonly from about 100-1000 mg/kg/day, preferably from about 150-700 mg/kg/day and most preferably from about 250-500 mg/kg/day. Preferably, therapeutically effective serum levels will be achieved by administering multiple doses each day. In cases of local administration or selective uptake, the effective local concentration of the drug may not be related to plasma concentration. One having skill in the art will be able to optimize therapeutically effective local dosages without undue experimentation.
When used as contraceptive agents in the female, the compositions of the invention can be evaluated for their effectiveness by any of a number of art accepted parameters including number of follicles, number of oocytes, number of transferrable embryos, number of pregnancies, the total dose administered and the treatment length. Similarly accepted criteria are available for evaluating the safety of a contraceptive agent. When used as contraceptive agents in the male, effectiveness can be adduced by decreased sperm count, sperm motility and the like. Additional criteria and methods for assessing the efficacy of a thiazolidinone-containing pharmaceutical composition, when used as a contraceptive agent or for another purpose, will be apparent to those of skill in the art.
The thiazolidinones can be incorporated into the pharmaceutical formulation as mixtures of diastereomers, mixtures of enantiomers or as stereochemically distinct compounds. The origin of the isomerism is the chirality of the carbons at positions 2 and 5 of the thiazolidinone ring structure (Formula I). For example in one preferred embodiment, the thiazolidinone component of the pharmaceutical composition is a mixture of cis and trans isomers. In another preferred embodiment, the mixture of cis and trans isomers is enriched in the cis isomer relative to the trans isomer. In a further preferred embodiment, the thiazolidinone is present as the substantially pure cis isomer.
The stereochemistry of the carbon atoms at positions 2 and 5 of the ring is yet another feature of the thiazolidinone constituent which can be varied. In a preferred embodiment, the thiazolidinone constituent is a mixture of the 2S, 5R and 5S, 2R isomers. In a more preferred embodiment, the thiazolidinone constituent is enriched in the 2S, 5R isomer. In still further preferred embodiments, the thiazolidinone constituent is substantially pure 2S, 5R.
In addition to the foregoing, the compounds of the invention are useful in vitro as unique tools for understanding the biological role of FSH, including the evaluation of the many factors thought to influence, and be influenced by, the production of FSH and the interaction of FSH with the FSH-R (e.g., the mechanism of FSH signal transduction/receptor activation). The present compounds are also useful in the development of other compounds that interact with the FSH-R, because the present compounds provide important structure-activity relationship (SAR) information that facilitate that development.
Compounds of the present invention that bind to the FSH receptor can be used as reagents for detecting FSH receptors on living cells, fixed cells, in biological fluids, in tissue homogenates, in purified, natural biological materials, etc. For example, by labelling such compounds, one can identify cells having FSH-R on their surfaces. In addition, based on their ability to bind the FSH receptor, compounds of the present invention can be used in in situ staining, FACS (fluorescence-activated cell sorting), western blotting, ELISA (enzyme-linked immunoadsorptive assay), etc. In addition, based on their ability to bind to the FSH receptor, compounds of the present invention can be used in receptor purification, or in purifying cells expressing FSH receptors on the cell surface (or inside permeabilized cells).
The compounds of the invention can also be utilized as commercial research reagents for various medical research and diagnostic uses. Such uses can include but are not limited to: (1) use as a calibration standard for quantitating the activities of candidate FSH antagonists in a variety of functional assays; (2) use as blocking reagents in random compound screening, i.e., in looking for new families of FSH receptor ligands, the compounds can be used to block recovery of the presently claimed FSH compounds; (3) use in the co-crystallization with FSH receptor, i.e., the compounds of the present invention will allow formation of crystals of the compound bound to the FSH receptor, enabling the determination of receptor/compound structure by x-ray crystallography; (4) other research and diagnostic applications wherein the FSH-receptor is preferably inactivated or such inactivation is conveniently calibrated against a known quantity of an FSH antagonist, and the like; (5) use in assays as probes for determining the expression of FSH receptors on the surface of cells; and (6) developing assays for detecting compounds which bind to the same site as the FSH receptor binding ligands.